1. Field of the Invention
The present invention relates generally to the field of medicine. More particularly, it concerns methods for delivering bioactive factors to bone.
2. Description of Related Art
Numerous pathological conditions are associated with abnormal bone cell function including osteoporosis, osteoarthritis, Paget's disease, osteohalisteresis, osteomalacia, periodontal disease, bone loss resulting from multiple myeloma and other forms of cancer, bone loss resulting from side effects of other medical treatment (such as steroids), and age-related loss of bone mass. Loss of bone mass in particular can lead to skeletal failure such that bone fractures can result from the minimal trauma of everyday life. Such fractures cause significant illness, or morbidity, inasmuch as there is insufficient repair or healing of the fractures.
Osteoporosis is the most common cause of bone loss and is a leading cause of disability in the elderly, particularly in elderly women. Osteoporosis is a progressive disease which results in the reduction of total bone mass and increased bone fragility. This often results in spontaneous fractures of load-bearing bones and the physical and mental deterioration characteristic of immobilizing injuries. The most widely accepted preventive agent for osteoporosis currently in use is estrogen therapy. However, systemic administration of estrogen is not a viable option in women at elevated risk for breast or endometrial cancers (estrogen dependent tumors) or for men (Cooper, 1994). In addition, recent studies have shown that estrogen replacement therapies (ERT's) have other deleterious side-effects, calling into question the long-term effects of these therapies.
Bisphosphonates have been effective inhibitors of osteoclastic bone resorption and have been used to advantage in treating osteoporosis (Parfitt el al., 1996). Bisphosphonates have been shown to increase trabecular bone volume and inhibit the decrease in cancellous bone mass in hindlimb unloaded rats compared to treated controls. However, the amount of cartilage in trabecular bone in these animals significantly increases, indicating that the modeling process is altered and mineralized cartilage fails to be resorbed and replaced by bone.
Vitamin D (1,25D), calcium and thiazide diuretics have also been used alone or in combination to prevent bone loss associated with corticosteroid treatment. The goal of such therapy is to improve calcium absorption and decrease urinary excretion of calcium thus, reversing secondary hyperparathyroidism (Joseph, 1994). Calcium supplements are widely used in managing established osteoporosis but there have been few satisfactory prospective studies of calcium supplementation on bone density or on the risk of future fractures (Cooper, 1994).
Bone damage, such as bone fractures, represents another common bone malady. Although repair, healing and augmentation of bone require a complex series of events that are not well defined, it is known that specific, naturally occurring factors are required to achieve these objectives. Such factors are released or migrate into the injured areas, and stimulate osteoblasts, chondrocytes, and odontoblasts in bone and cartilage to stimulate matrix formation and remodeling of the wounded area (ten Dijke et al., 1989).
New bone can be formed by three basic mechanisms: osteogenesis, osteoconduction and osteoinduction. Cancellous bone and marrow grafts provide viable cells for such processes. Transforming growth factor-beta (TGF-β) has been shown to stimulate proliferation and matrix synthesis of osteoblastic cells (Centrella et al., 1987) and to inhibit the formation and activity of osteoclastic cells (Chenu et al. 1988). Members of the bone morphogenetic protein family have been shown to be useful for induction of cartilage and bone formation. For example, BMP-2 has been shown to be able to induce the formation of new cartilage and/or bone tissue (U.S. Pat. No. 5,013,649).
Weightlessness during spaceflight has also been a cause of bone loss. Countermeasures for such bone loss have been of the skeletal stress type, such as cycling, simulated running, and rowing (Baldwin el al., 1996). Studies show that exercise-induced skeletal stress serves to maintain and increase osteoblastic activity. However, these methods alone are insufficient to prevent bone volume losses, primarily because it is not possible to generate forces of equal magnitude to those encountered on Earth (McCarthy et al., 2000; Baldwin et al., 1996). Electrical stimulation of selected muscle groups in hindlimb unloaded rats also increases osteoblast activity and osteoid surfaces, but does not prevent decrease in trabecular bone volume or metaphyseal apposition rate (Zerath et al., 1995). Bisphosphonates such as alendronate minimize bone loss during unloading by inhibiting osteoclastic bone resorption, but do not prevent the unloading-induced suppression of bone formation (Bikle et al., 1994). Thus, antiresorbing agents are not ideal countermeasures to bone loss when the primary defect is reduced bone formation (McCarthy et al, 2000).
Growth hormone (GH) treatment of hypophysectomized rats has been shown to increase bone mass independent of whether the animals are loaded or unloaded. However, unloaded animals still show lower bone mass relative to treated animals for the same treatment protocol. Pharmacological doses of GH of 500 μg/ml also failed to mediate skeletal defects in hypophysectomized rats in response to hindlimb unloading, including decreased trabecular bone volume and cortical bone apposition rate (Halloran et al., 1995). Although systemic factors such as GH and 1,25D may modulate the response of bone to unloading, factors that locally regulate bone growth may have greater utility as countermeasure molecules to prevent bone loss.
While the above described countermeasures have been successful in minimizing to some extent the morbidity associated with abnormal bone cell function, the efficacy of such treatments is limited by the ability to appropriately deliver the active ingredient to the site where needed. In addition, most of these treatments have serious side effects when administered systemically.
Therefore, the need for site specific targeting of therapeutic agents has been felt. However, site-specific targeting requires quantitatively distinct receptors that are unique to bone. A few researchers have demonstrated that the inorganic component of bone which is comprised of hydroxyapatite (HAp), occurs normally only in hard tissues. A bisphosphonate, methylene bisphosphonate (MBP), is known for its predilection to bone sites undergoing remodeling, has been used in combination with Technetium-99 m (99mTc) as a non-therapeutic diagnostic imaging tool in the study of bone pathology (Davis and Jones, 1976; Lantto et al., 1987; Cronhjort et al., 1999). MBP has been studied as a bone matrix targeting moiety for osteotropic drug delivery. Fujisaki, et al., (1995; 1996), conjugated various model materials and pro-drug candidates to MBP and demonstrated their targeting efficacy in vivo. Estradiol conjugated to MBP was taken up in bone and then released from MBP either by enzymatic or chemical hydrolysis of the ester conjugation linkage. Uludag, et al., (2000a; 2000b), demonstrated the osteotropic delivery of model proteins conjugated to MBP by similar chemistry.
In addition, several bone non-collagenous proteins, such as osteopontin and bone sialoprotein, are known to contain amino residue sequences that bind specifically to HAp (Nagata et al., 1991). Fujisawa et al., determined that a six-residue aspartic acid oligopeptide (Asp6) preferentially binds to the calcified matrix in vivo (Kasugai et al., 2000) and that this targeting ligand can deliver an estradiol pro-drug in vivo (Yokogawa et al., 2000).
Prior approaches for targeting bone that use simple molecules conjugated to bone-targeting ligands that preferentially accumulated in bone have serious drawbacks. First, conjugated molecules are systemically exposed and, therefore, are subject to rapid elimination or can have action on sites other than bone. Second, conjugation of the bone-targeting ligands to the therapeutic molecules can adversely alter their therapeutic activity. Lastly, release of the active molecule from the targeting ligand, and its subsequent activity, is dependent on the degradation kinetics of the conjugation linkage.
Nonetheless, the aging global population translates to ever-increasing demand for orthopedic countermeasures to skeletal deterioration resulting from the increasing fragility of skeletal structures with age. In addition, there is an acute need to find effective countermeasures for other bone conditions and ailments. There is, therefore, a great need in the art for novel therapeutic compositions that can be used to deliver therapeutic agents to targets in the bone for effective treatment of bone-associated maladies.